Cancer Res Treat.  2015 Oct;47(4):913-920. 10.4143/crt.2014.057.

A Novel, Potent, Small Molecule AKT Inhibitor Exhibits Efficacy against Lung Cancer Cells In Vitro

• Dinavahi S
• Prasanna R
• Dharmarajan S
• Perumal Y
• Viswanadha S

• KMID: 2403412
• DOI: http://doi.org/10.4143/crt.2014.057

Abstract

PURPOSE
Anomalies of Akt regulation, including overexpression in lung cancer, impart resistance to conventional chemotherapy and radiation, thereby implicating this kinase as a therapeutic intervention point. A novel scaffold of Akt inhibitors was developed through virtual screening of chemical databases available at Birla Institute of Technology and Science, Pilani, Hyderabad, based on docking studies using Maestro. A benzothienopyrimidine derivative (BIA-6) was identified as a potential lead molecule that inhibited Akt1 enzyme activity with an IC50 of 256 nM.
MATERIALS AND METHODS
BIA-6 was tested for in vitro Akt1 inhibition using a fluorescence resonance energy transfer kit. Anti-proliferative activity was tested in NCI-H460, A549, NCI-H1975, and NCI-H2170 cell lines. The effect of the compound on p-Akt (S473) was estimated.
RESULTS
BIA-6 allosterically caused a dose dependent reduction of growth of cell lines with a half maximal growth inhibition (GI50) range of 0.49 muM to 6.6 muM. Cell cycle analysis indicated that BIA-6 caused a G1 phase arrest at < 100 nM but led to apoptosis at higher doses. BIA-6 also exhibited synergism with standard chemotherapeutic agents.
CONCLUSION
BIA-6 is a novel, allosteric Akt inhibitor with potent anti-cancer activity in lung cancer cell lines, that effectively blocks the phosphoinositide-3 kinase/Akt pathway with a high margin selectivity towards normal cells.

Keyword

Akt1; PI3KCA; Non-small-cell lung carcinoma; BIA-6; Apoptosis; Drug synergism

MeSH Terms

Apoptosis
Carcinoma, Non-Small-Cell Lung
Cell Cycle
Cell Line
Databases, Chemical
Drug Synergism
Drug Therapy
Fluorescence Resonance Energy Transfer
G1 Phase
Inhibitory Concentration 50
Lung Neoplasms*
Lung*
Mass Screening
Phosphotransferases
Phosphotransferases

Figure

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  Fig. 1. (A) Expression of p-Akt in various lung cancer cell lines. Cells were treated with BIA-6 for 4 hours, lysed and p-Akt expression was quantified by Western blot. Representation of lanes: 1, A549; 2, H460; 3, H1975; 4, H2170; 5, HTEpiC; and 6, MDA-MB-231. MDA-MB-231 was taken as a negative control cell line. (B) Markush structure for BIA-6.

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  Fig. 2. Effect of BIA-6 on cellular proliferation in lung cancer cells (A) and normal cells (B). HUVEC, human umbilical vascular endothelial cells; HTEpiC, human tracheal epithelial cell.

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  Fig. 3. BIA-6 induces apoptosis in lung cancer cell lines. One-way ANOVA with Dunnett’s as post hoc analysis compared to dimethyl sulfoxide (DMSO) control. RFU, relative fluorescence unit. *p < 0.05 , **p < 0.01, ***p < 0.001.

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  Fig. 4. BIA-6 arrests the cell cycle in G1 phase in lung cancer cell lines. Cells were incubated with 100 nM BIA-6 (+) or dimethyl sulfoxide (DMSO) (–) for 72 hours before cell cycle analysis. One-way ANOVA with Dunnett’s as post hoc analysis compared to DMSO control. *p < 0.05, **p < 0.01.

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  Fig. 5. Cellular efficacy is due to the inhibition of Akt pathway. Cells were treated with BIA-6 for 4 hours, lysed and p-Akt was estimated by Western blot. The levels of p-Akt were quantified at 100 nM and 1,000 nM concentration in lung cancer cell lines. The levels of p-Akt were quantified at 100 nM concentration in lung cancer cell lines.

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  Fig. 6. Combination of BIA-6 with gemcitabine/docetaxel in lung cancer cell lines. The concentrations of the drugs are as follows: B, BIA-6, 1,000 nM; G, gemcitabine, 100 nM; D, docetaxel, 100 nM. Values in the graph indicate the combination indices at the respective concentrations. Combination index < 1 indicates synergism.

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  Appendix 1. Effect of BIA-6 on cellular proliferation in breast and colon cancer.

Reference

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